The biosynthesis of methionine-enkephalin and leucine-enkephalin, pentapeptides which are endogenous ligands for the opiate receptor, is under investigation. Messenger RNA obtained from bovine adrenal medulla, a rich source of enkephalins, has been isolated and used as a template for the synthesis of the primary enkephalin gene product in cell-free translation systems. The synthesis of an enkephalin-containing putative precursor protein was demonstrated by the liberation of enkephalin sequences from protein by specific proteolysis, followed by immunoprecipitation and high-performance liquid chromatography. The moleculr weight of the enkephalin-containing protein is probably 1130-32,000, while the messenger RNA coding for this protein is heterogeneous and consists of possibly two fractions of apparent size 4750 and 3000 nucleotides. Monoclonal antibodies and conventional antibodies to enkephalin precursor protein are being prepared, and a nucleic acid probe for enkephalin messenger RNA is being utilized to identify and quantitate enkephalin messenger RNA. Using the methodology above, regulation of enkephalin precursor gene expression is under investigation